Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons

Background: Mutations in leucine-rich repeat kinase 2 (LRRK2) contribute to both familial and idiopathic forms of Parkinson's disease (PD). Neuroinflammation is a key event in neurodegeneration and aging, and there is mounting evidence of LRRK2 involvement in inflammatory pathways. In a previous study, we described an alteration of the inflammatory response in dermal fibroblasts from PD patients expressing the G2019S and R1441G mutations in LRRK2. Methods: Taking advantage of cellular reprogramming, we generated induced pluripotent stem cell (iPSC) lines and neurons thereafter, harboring LRRK2G2019S and LRRK2R1441G mutations. We used gene silencing and functional reporter assays to characterize the effect of the mutations. We examined the temporal profile of TNF alpha-induced changes in proteins of the NF-kappa B pathway and optimized western blot analysis to capture alpha-synuclein dynamics. The effects of the mutations and interventions were analyzed by two-way ANOVA tests with respect to corresponding controls. Results: LRRK2 silencing decreased alpha-synuclein protein levels in mutated neurons and modified NF-kappa B transcriptional targets, such as PTGS2 (COX-2) and TNFAIP3 (A20). We next tested whether NF-kappa B and alpha-synuclein pathways converged and found that TNF alpha modulated alpha-synuclein levels, although we could not detect an effect of LRRK2 mutations, partly because of the individual variability. Nevertheless, we confirmed NF-kappa B dysregulation in mutated neurons, as shown by a protracted recovery of I kappa B alpha and a clear impairment in p65 nuclear translocation in the LRRK2 mutants. Conclusions: Altogether, our results show that LRRK2 mutations affect alpha-synuclein regulation and impair NF-kappa B canonical signaling in iPSC-derived neurons. TNFa modulated alpha-synuclein proteostasis but was not modified by the LRRK2 mutations in this paradigm. These results strengthen the link between LRRK2 and the innate immunity system underscoring the involvement of inflammatory pathways in the neurodegenerative process in PDThis study is funded by grants from the Spanish Ministry of Economy and Competitiveness (MINECO), Fondo de Investigaciones Sanitarias PI15/00486, the European Commission FP7 Health -278871, and the Joint Program in Neurodegenerative Diseases AC 14/0041 (DAMNDPATHS) to RSP

Fast and efficient neural conversion of human hematopoietic cells

Neurons obtained directly from human somatic cells hold great promise for disease modeling and drug screening. Available protocols rely on overexpression of transcription factors using integrative vectors and are often slow, complex, and inefficient. We report a fast and efficient approach for generating induced neural cells (iNCs) directly from human hematopoietic cells using Sendai virus. Upon SOX2 and c-MYC expression, CD133-positive cord blood cells rapidly adopt a neuroepithelial morphology and exhibit high expansion capacity. Under defined neurogenic culture conditions, they express mature neuronal markers and fire spontaneous action potentials that can be modulated with neurotransmitters. SOX2 and c-MYC are also sufficient to convert peripheral blood mononuclear cells into iNCs. However, the conversion process is less efficient and resulting iNCs have limited expansion capacity and electrophysiological activity upon differentiation. Our study demonstrates rapid and efficient generation of iNCs from hematopoietic cells while underscoring the impact of target cells on conversion efficiency

Characterization of promising potato clones (solanum tuberosum l. subspecies andigena) for starch extraction

Colombia has been overproducing potatoes with around 18% remaining unmarketable, constituting a potential alternative use in obtaining native starch for the food industry. To this end, 17 promising potato clones from the Programa de Mejoramiento Genetico of Corpoica were characterized for agronomic variables, as well as physicochemical variables for the tubers. The results were analyzed with descriptive statistics, Pearson correlation and cluster analysis. Clone codified 36 was selected as having potential for the extraction of native starch, showing agronomically: a plant height of 75 cm, a green and undulating stalk, moderate deep purple and white blooming, semi-stellate-shaped corolla, green globe-shaped berries, oval tubers with a creamy white peel color and dotted with purple spots; physicochemically the tubers had: 1.090 specific gravity, 23.12% dry matter, 16.82% starch (22% amylose and 78% amylopectin), 0.122% reducing sugars, 2% protein, 0.82% ash and a pH of 5.69. The extracted starch showed a yield of 53.66% with 19.32% moisture

Fast and efficient neural conversion of human hematopoietic cells

Neurons obtained directly from human somatic cells hold great promise for disease modeling and drug screening. Available protocols rely on overexpression of transcription factors using integrative vectors and are often slow, complex, and inefficient. We report a fast and efficient approach for generating induced neural cells (iNCs) directly from human hematopoietic cells using Sendai virus. Upon SOX2 and c-MYC expression, CD133-positive cord blood cells rapidly adopt a neuroepithelial morphology and exhibit high expansion capacity. Under defined neurogenic culture conditions, they express mature neuronal markers and fire spontaneous action potentials that can be modulated with neurotransmitters. SOX2 and c-MYC are also sufficient to convert peripheral blood mononuclear cells into iNCs. However, the conversion process is less efficient and resulting iNCs have limited expansion capacity and electrophysiological activity upon differentiation. Our study demonstrates rapid and efficient generation of iNCs from hematopoietic cells while underscoring the impact of target cells on conversion efficiency

LRRK2 Expression Is Deregulated in Fibroblasts and Neurons from Parkinson Patients with Mutations in PINK1

Mutations in PINK1 (PARK6), a serine/threonine kinase involved in mitochondrial homeostasis, are associated with early onset Parkinson’s disease. Fibroblasts from Parkinson’s disease patients with compound heterozygous mutations in exon 7 (c.1488 + 1G > A; c.1252_1488del) showed no apparent signs of mitochondrial impairment. To elucidate changes primarily caused by lack of functional PINK1, we over-expressed wild-type PINK1, which induced a significant downregulation of LRRK2 (PARK8). Indeed, we found that LRRK2 protein basal levels were significantly higher in the mutant PINK1 fibroblasts. To examine the interaction between the two PARK genes in a disease-relevant cell context, we generated induced pluripotent stem cell (iPSC) lines from mutant, carrier and control fibroblasts by lentiviral-mediated re-programming. Efficiency of neural induction and dopamine differentiation using a floor-plate induction protocol was similar in all genotypes. As observed in fibroblasts, PINK1 mutant neurons showed increased LRRK2 expression both at the RNA and protein level and transient over-expression of wild-type PINK1 efficiently downregulated LRRK2 levels. Additionally, we confirmed a dysregulation of LRRK2 expression in fibroblasts from patients with a different homozygous mutation in PINK1 exon 4, c.926G > A (G309D). Thus, our results identify a novel role of PINK1 modulating the levels of LRRK2 in Parkinson’s disease fibroblasts and neurons, suggest a convergent pathway for these PARK genes, and broaden the role of LRRK2 in the pathogenesis of Parkinson’s disease.This study is funded by grants from the European Commission, CORDIS FP7, HEALTH-2, 278871 (DDPDGENES), and the Joint Program in Neurodegenerative Diseases (DAMNDPATHS) to RSP. JPB is the recipient of grants from MINECO (SAF2013-41177-R) and the NIH/NIDA (1R21DA037678-01). GAu is the recipient of grants from DFG (AU96/10-1) and the German National Genome Research Network NGFNplus (Neuron-Parkinson-subproject 7).Peer reviewe

LRRK2 Expression Is Deregulated in Fibroblasts and Neurons from Parkinson Patients with Mutations in PINK1.

Mutations in PINK1 (PARK6), a serine/threonine kinase involved in mitochondrial homeostasis, are associated with early onset Parkinson's disease. Fibroblasts from Parkinson's disease patients with compound heterozygous mutations in exon 7 (c.1488 + 1G > A; c.1252_1488del) showed no apparent signs of mitochondrial impairment. To elucidate changes primarily caused by lack of functional PINK1, we over-expressed wild-type PINK1, which induced a significant downregulation of LRRK2 (PARK8). Indeed, we found that LRRK2 protein basal levels were significantly higher in the mutant PINK1 fibroblasts. To examine the interaction between the two PARK genes in a disease-relevant cell context, we generated induced pluripotent stem cell (iPSC) lines from mutant, carrier and control fibroblasts by lentiviral-mediated re-programming. Efficiency of neural induction and dopamine differentiation using a floor-plate induction protocol was similar in all genotypes. As observed in fibroblasts, PINK1 mutant neurons showed increased LRRK2 expression both at the RNA and protein level and transient over-expression of wild-type PINK1 efficiently downregulated LRRK2 levels. Additionally, we confirmed a dysregulation of LRRK2 expression in fibroblasts from patients with a different homozygous mutation in PINK1 exon 4, c.926G > A (G309D). Thus, our results identify a novel role of PINK1 modulating the levels of LRRK2 in Parkinson's disease fibroblasts and neurons, suggest a convergent pathway for these PARK genes, and broaden the role of LRRK2 in the pathogenesis of Parkinson's disease

The relationship between coping style and psychological distress in people with head and neck cancer: a systematic review

Objective Individuals diagnosed with head and neck cancer (HNC) are at an elevated risk of experiencing psychological distress and a reduced quality of life. The aim of this review was to systematically examine and assess the quality of empirical evidence on the associations between coping mechanisms and psychological distress among people with HNC. Methods CINAHL, MEDLINE, PsycINFO, EMBASE and Web of Science were accessed to conduct this review. Studies were included if they used reliable and valid measures to investigate the relationship between coping style and psychological distress. Study quality was assessed and rated according to pre-set criteria, and showed variability in relation to selection methods. Results 12 studies (nine cross-sectional and three prospective designs) involving 1281 patients were reviewed. There was considerable heterogeneity in study samples and coping measures. Moderate to large associations between disengagement coping mechanisms (e.g. avoidance) and psychological distress were observed. Engagement coping strategies (e.g. direct action) were not consistently associated with psychological distress across studies. Conclusions Several studies observed a significant relationship between coping styles aimed at disengaging and distancing from cancer and increased psychological distress. To understand directionality of these associations and further develop an understanding of temporal features of the relationship between coping styles and distress, longitudinal designs could be used in future research